564 AMPHIBIAN AND REPTILE DISEASES Herpetological Review, 2017, 48(3), 564–568. © 2017 by Society for the Study of Amphibians and Reptiles Swabbing for Batrachochytrium salamandrivorans on Wild Rough-skinned Newts (Taricha granulosa) and Pet-Traded Amphibians on Southern Vancouver Island, British Columbia, Canada Batrachochytrium salamandrivorans (Bsal) is a fungal urban and rural areas, known to researchers to have contained pathogen that causes chytridiomycosis in salamanders and is Rough-skinned Newts in the past. To capture wild Rough- similar to the more widely known B. dendrobatidis (Bd) that is skinned Newts, minnow traps were deployed overnight and believed to have been responsible for a global decline in over 200 checked in the morning. Dip nets were also used at some sites. amphibian species (Skerratt et al. 2007). Bsal is known to infect Captured Rough-skinned Newts were swabbed following the the skin of salamanders, causing skin lesions, anorexia, apathy, AmphibiaWeb “chytrid swabbing protocol” (Briggs NIH Research ataxia, and death (Martel et al. 2013). Bsal has been reported to Group 2009) and then released. To avoid the spread of diseases have caused localized mortality of wild and captive salamander from sample sites, the “British Columbia hygiene protocol for species in Europe (Spitzen-van der Sluijs et al. 2013; Martel et field staff working in aquatic environments” (Ecosystems Branch, al. 2014; Sabino-Pinto et al. 2015). It has been hypothesized that British Columbia Ministry of Environment 2008) was followed. Bsal was spread to Europe on human-transported salamanders In addition to testing wild Rough-skinned Newts for this originating in Asia, later spreading to wild populations (Martel study, researchers tested for the presence of Bsal on captive et al. 2014). There is a concern that Bsal could impact native pet traded salamanders. The intention was to help determine salamanders in North America through this introductive if pet traded salamanders could be acting as vectors for Bsal. pathway (Gray et al. 2015; Stephen et al. 2015). In laboratory To locate pet-traded salamanders, over 30 pet stores were challenge experiments, Martel et al. (2014) reported Rough- contacted across British Columbia by telephone and asked skinned Newts (Taricha granulosa), a salamander endemic to about their salamander inventory, and in some but not all cases, the Pacific Northwest of North America, died within a few days of their anuran inventory. All but one pet store indicated that they Bsal exposure. As a result of the risk of Bsal to native amphibians did not have salamanders, but many indicated that they sold in North America, in 2015 the Pet Industry Joint Advisory various species of anurans. Researchers did not further verify Council (PIJAC) of Canada, promoted a voluntary moratorium pet store information by visiting each store, due to cost and time on the importation of Fire-bellied Newts (Cynops orientalis) and limitations for this study. Paddle-tailed Newts (Pachytriton labiatus) into Canada (PIJAC By the end of the sampling period, 15 captive salamanders 2015); in 2016 the US Fish and Wildlife Service instated an interim had been swabbed by researchers. One store in Victoria (Pet Store prohibition on the importation of 20 genera of salamanders into 1) was found to have 13 Fire-bellied Newts (Cynops orientalis) the United States (US Fish and Wildlife Service 2016); and in and 1 Axolotl (Ambystoma mexicanum), which were all swabbed May 2017 Canada instated a one-year import restriction on all (Table 1). One Barred Tiger Salamander (Ambystoma mavortium) salamander species within the order Caudata (Environment and was also opportunistically swabbed after being confiscated by Climate Change Canada 2017). the British Columbia Ministry of Environment from a pet store This study is the first field-based effort to test for the in the Lower Mainland, British Columbia (Pet Store 3; Table 1). presence of Bsal on wild Rough-skinned Newts and pet-traded Barred Tiger Salamanders are listed under Schedule A of the salamanders on southern Vancouver Island, British Columbia, British Columbia Wildlife Act as “Wildlife” which are illegal to Canada. Southern Vancouver Island was the chosen study region possess in captivity. While visiting Pet Store 1 (to initially swab to test wild salamanders for two reasons. Firstly, it is part of the salamanders) and Pet Store 2 in Gibsons, British Columbia Rough-skinned Newt’s native range, and secondly it has been (opportunistic visit by researcher while travelling), 15 captive indicated as a high vulnerability zone for Bsal in North America anurans were additionally swabbed (Table 1). due to occurrence of known-susceptible species, suitable habitat Swabs from wild Rough-skinned Newts and from captive conditions, and import trade centers (Yap et al. 2015). pet-store salamanders and anurans were sent to the BC Ministry From May–July 2016, nine water bodies, small lakes, or ponds of Agriculture’s Animal Health Centre in Abbotsford, British with permanent water, were sampled on southern Vancouver Columbia. Swabs from Rough-skinned Newts were tested for Island for Rough-skinned Newts (Fig. 1). Sites were chosen from Bsal, and the swabs from pet store salamanders and anurans were tested for both Bsal and Bd using quantitative polymerase PURNIMA GOVINDARAJULU chain reaction (qPCR) analyses (Bsal: Blooi et al. 2013; Bd: Boyle British Columbia Ministry of Environment, Victoria, BC V8T 5J9, et al. 2004). Samples were run as single reactions and both Bd Canada; e-mail: [email protected] PCR and Bsal PCR were run independently. Exogenous internal ELYSE MATTHEWS controls, negative extraction controls, no-template controls, and British Columbia Ministry of Environment, Victoria, BC V8T 5J9, positive controls were used for both Bd and Bsal PCRs. Previously, Canada; e-mail: [email protected] KRISTIINA OVASKA the BC Ministry of Agriculture’s Animal Health laboratory had Habitat Acquisition Trust, Victoria, BC V8W 1E4, Canada participated in a ring test for Bd and Bsal, organized by the e-mail: [email protected] Bsal Task Force Diagnostic Working Group (H. Snyman, pers, comm., Animal Health Centre, British Columbia Ministry of Herpetological Review 48(3), 2017 AMPHIBIAN AND REPTILE DISEASES 565 that have been undergoing an importation ban. One importer in Vancouver, British Columbia was interviewed by researchers over the phone, and stated that their business had stopped ordering salamanders after receiving a call from PIJAC about the voluntary moratorium. Prior to this call from PIJAC, the wholesaler had been ordering 100–200 salamanders on a bi-weekly basis to supply retailers across British Columbia (Vancouver wholesaler, pers. comm., 2016). This is an indication of the application of PIJAC’s past moratorium, and the importance of working with wholesalers and industry representatives in the future to address concerns about Bsal. While this importer’s account is important to understand the impacts of PIJAC’s moratorium and the US prohibition, a larger survey, directed towards large retail and independent pet stores, wholesalers, online and private traders, and hobbyists, is warranted. It would be valuable for future studies to gather information across the spectrum of stakeholders, to get a sense of their knowledge and experiences with PIJAC’s moratorium, the US import prohibitions, and the recent Canadian importation FIG. 1. Sample sites for Batrachochytrium salamandrivorans on wild restrictions. It is important to capture such variation when Rough-skinned Newts (Taricha granulosa) on southern Vancouver making conclusions on the impacts of these moratoriums and Island, British Columbia, Canada. prohibitions. Even though this study was intended to test for Bsal, Agriculture, November 2016; Bsal Task Force 2016). This ring test opportunistic testing of captive, pet store amphibians for Bd was intended to test for standardization of laboratory methods was also possible. As a result of this testing, we found that and results across North American laboratories in Canada and Bd was detected in one of 15 swabbed anurans. In British the US. Columbia, chytridiomycosis caused by Bd is a reportable Bsal was not detected by qPCR analyses of swabs from the disease. Unfortunately, the infected anuran had been sold before 82 wild Rough-skinned Newts and 15 captive salamanders. analyses of the samples were completed; therefore, a second Additionally, Bd was not detected from samples of the captive test and proceeding quarantine measures were not taken. The salamanders tested (wild salamanders were not tested). From detection of Bd on this individual indicates that Bd of unknown samples of the 15 captive anurans, Bsal was not detected (Table and potentially virulent strains could be transferred by the pet 1), however, Bd was detected from one captive Ornate Pacman trade into British Columbia. Introduced American Bullfrogs Frog (Ceratophrys ornata) (Table 1). swabbed in the wild, have been found to be carriers of strains of In our preliminary surveys, we found no evidence that Bsal Bd on southern Vancouver Island (Leblanc 2012). It is possible had infected wild Rough-skinned Newts on southern Vancouver that if other non-native anurans are introduced, they could also Island. However, it is recommended that future studies increase be carriers of strains of non-native chytrid fungi. the sample size and number of sites to allow for greater Bsal was not detected in this study on swabbed captive confidence. In addition, expanding sampling across a wider area anurans. Recently however, Iwanowicz et al. (2017) reported of the Rough-skinned Newt’s native range would be desirable, laboratory detections of Bsal from a Rough-skinned Newt and including the Lower Fraser Valley where newt populations are American Bullfrog (Lithobates catesbeianus) in Oregon, USA, embedded within populated and highly modified landscapes. but the result could not be confirmed with additional testing, Bsal was not detected in any of the captive salamanders which supported either a contaminated sample or detection of sampled. However, the small sample size (N = 15) and small a Bsal -like organism.